Laboratory Studies
Stool examination to visualize the ova or adult worms is the diagnostic method of choice.
The eggs of heterophyid species are smaller than eggs of other intestinal flukes. Modified Kato thick method has a higher sensitivity than formalin-ether/ethyl acetate concentration technique (31% vs 13.6%) in detecting heterophyid eggs. [38] Heterophyid species are difficult to distinguish and may also be confused with Clonorchis and Opisthorchis eggs. In areas endemic for liver flukes, heterophyid eggs have been referred to as "opisthorchid-like eggs."
Other laboratory findings include anemia and eosinophilia.
Serologic tests have limited application; however, for certain combinations of pathogens and their available diagnostic testing, serodiagnosis may be helpful, as in the case of F buski infections. [41]
Chronic fascioliasis is generally evaluated with fecal egg counting after concentration of the eggs in the stool sample via a zinc sulphate floatation method. However, using the sedimentation technique to concentrate the eggs is said to improve sensitivity. A F hepatica coproantigen enzyme-linked immunoassay (ELISA) has been introduced and studied in cattle and sheep. It more accurately reflects the presence of flukes in the host bile ducts in late prepatent infections and clearance of the flukes after treatment. [42] It can probably be used in humans in the future.
Urine assay, particularly of O viverrini excretory-secretory (ES) antigens in urine, has been used to detect O viverrini in Thailand. It was found easier to use and more sensitive than the traditional ethyl-acetate concentration technique. [43]
Diagnosing O viverrini infection via conventional stool examination is difficult, both because the infection may decrease in intensity after repeated treatments under control programs in endemic areas and because of the presence of coinfections with intestinal flukes. Thus, one study has examined a coproantigen sandwich ELISA using recombinant O viverrini cathepsin F (rOv-CF) that uses chicken immunoglobulin Y (IgY) raised against rOv-CF in combination with rabbit immunoglobulin G (IgG) antibody to the somatic O viverrini antigens. This test showed a sensitivity and specificity of 93.3% and 76.7%, respectively, in the detection of opisthorchiasis. The investigators found that it had a positive predictive value (PPV) and negative predictive value (NPV) of 66.7% and 95.2%, respectively, making it a promising test in endemic areas. [44]
The current criterion standard of diagnosis is the formalin ethyl-acetate concentration technique (FECT), performed with fecal samples. However, this test has difficulty detecting light O viverrini infections since the eggs may be confused with eggs of other minute intestinal flukes in stool. [43]
Other Tests
Merthiolate, iodine, formalin method
The merthiolate, iodine, formalin (MIF) method is used to detect intestinal fluke parasites.
The MIF method was established early as a versatile and accurate technique for identifying intestinal protozoa in stool and fecal samples. [45] The technique simultaneously preserves and stains stool specimens, which can then be examined with direct smear techniques. [46]
Following the development of a concentrated MIF technique, the sensitivity of positively identifying F buski, Heterophyes species, and Echinostoma species in stool specimens was increased. [47] This newly concentrated MIF technique involves the application of a concentration step to the stool specimen before preservation in MIF solution.
Polymerase chain reaction
Various polymerase chain reaction (PCR) methods have shown potential in detecting intestinal fluke parasites. These methods take advantage of the different types of DNA nucleotide sequence variations demonstrated by the different species of parasites within a particular genus. [48]
In Thailand, Lamanignao et al developed a PCR-based technique to detect and discriminate between O viverrini and Haplorchis taichui. O viverrini and heterophyids have similar egg morphology, so an experienced microscopist is usually required for distinction. Both parasites have similar intermediate and definite hosts and may exist as coinfections. [49] More importantly, chronic O viverrini infection is associated with the development of cholangiocarcinoma. [50] Although both can be treated with praziquantel, Opisthorchis infection requires two days of treatment.
Likewise, the difficulty of diagnosing parasitic pathogens led Won et al to develop a multiplex qPCR to detect the presence of eight major intestinal parasites known to cause gastroenteritis in the Korean population. This includes three trematodes (C sinensis, M yokogawai, G seoi). Analysis included 123 stool samples, with the multiplex qPCR exhibiting 100% sensitivity (95% CI; 80.5-100) and 100% specificity (95% CI; 96.58-100). [51]
Polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) [17] and simple sequence repeat anchored PCR [52] have been reported to be useful in distinguishing among species of the Metagonimus genus (including M yokogawai). These methodologies are based on differences in restriction fragment length polymorphisms and simple sequence repeats among the species. Information derived from RFLP involving specific sites in ribosomal RNA and mitochondrial cytochrome oxidase I (mtCOI) genes may help to differentiate M yokogawai from other Metagonimus species. [53]
Six species of the genus Heterophyidae were reported to be distinguished with PCR assays developed based on variations in rDNA polymorphisms among the species. [54]
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Life cycle of Fasciolopsis buski. Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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The life cycle of Fasciolopsis. Immature eggs are discharged into the intestine and stool and become embryonated in water. The eggs then release miracidia, which invade a suitable snail intermediate host, in which the parasites undergo several developmental stages (sporocysts, rediae, cercariae). The cercariae are released from the snail and encyst as metacercariae on aquatic plants, which are eaten by mammalian hosts (humans and pigs), who become infected. After ingestion, the metacercariae excyst in the duodenum and attach to the intestinal wall, where they develop into adult flukes (20-75 mm X 8-20 mm) in approximately 3 months and attach to the intestinal wall of the mammalian hosts. The adults have a life span of about one year. Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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Egg of Fasciolopsis buski. Images reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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Adult fluke of Fasciolopsis buski. Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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The life cycle of Heterophyes. The adult parasites release embryonated eggs (each with a fully developed miracidium), which are then passed in the host's feces. After ingestion by a suitable snail (first intermediate host), the eggs hatch and release miracidia, which penetrate the snail's intestine. Snails of the genera Cerithidea and Pirenella are important hosts in Asia and the Middle East, respectively. The miracidia undergo several developmental stages in the snail (sporocysts, rediae, cercariae). Many cercariae are produced from each redia. The cercariae are released from the snail and encyst as metacercariae in the tissues of a suitable freshwater or brackish-water fish (second intermediate host). The definitive host becomes infected by ingesting undercooked or salted fish that contains metacercariae. After ingestion, the metacercariae excyst, attach to the mucosa of the small intestine, and mature into adults (measuring 1-1.7 mm X 0.3-0.4 mm). Heterophyes heterophyes infects humans, various fish-eating mammals (eg, cats, dogs), and birds. Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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Life cycle of Metagonimus. The adult parasites release fully embryonated eggs (each with a fully developed miracidium), which are then passed in the host's feces. After ingestion by a suitable snail (first intermediate host), the eggs hatch and release miracidia, which penetrate the snail's intestine. Snails of the genus Semisulcospira are the most common intermediate host for Metagonimus yokogawai. The miracidia undergo several developmental stages in the snail (sporocysts, rediae, cercariae). Many cercariae are produced from each redia. The cercariae are released from the snail and encyst as metacercariae in the tissues of a suitable freshwater or brackish-water fish (second intermediate host). The definitive host becomes infected by ingesting undercooked or salted fish that contains metacercariae. After ingestion, the metacercariae excyst, attach to the mucosa of the small intestine, and mature into adults (measuring 1-2.5 mm X 0.4-0.75 mm). M yokogawai infects humans, fish-eating mammals (eg, cats, dogs), and birds. Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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Various animals may be definitive hosts for different Echinostoma species, such as aquatic birds, carnivores, rodents, and humans. Unembryonated eggs are passed in stool (1), and development occurs in the water (2). The miracidium takes an average of 10 days to mature and then hatches (3), penetrating the first intermediate host, a snail (4). Snails, in general, serve as the first intermediate host. The intramolluscan stages are as follows: sporocyst (4a); rediae (4b); and cercariae (4c). Cercariae may then encyst as metacercariae in the same first intermediate host or leave to penetrate a new second intermediate host (5). Several animals may become the second intermediate host, such as other snails, bivalves, fish, and tadpoles. The definitive host gets infected after eating infected second intermediate hosts (6). The metacercariae excyst in the duodenum (7). Adults then live in the small intestine (8). Image reproduced from the Division of Parasitic Diseases, Centers for Disease Control and Prevention (CDC), Atlanta, GA.
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Magnified image of Echinostoma spp. egg. Echinostoma eggs resemble F busk and Fasciola eggs, with the latter two only bigger in size. Specimen slides courtesy of the University of the Philippines - College of Public Health, Department of Parasitology.
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Low power magnification (arrow) of a Heterophyid egg. Specimen slides courtesy of the University of the Philippines - College of Public Health, Department of Parasitology.
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Magnified view of a Heterophyid egg. Specimen slides courtesy of the University of the Philippines - College of Public Health, Department of Parasitology.
Tables
Infection |
Source* |
Geographic Distribution** |
|---|---|---|
Fasciolopsiasis |
Freshwater plants (water caltrop, water chestnut) |
Asia and Indian subcontinent, especially in areas where humans raise pigs and consume freshwater plants (as of December 8, 2017) |
Echinostomiasis |
Tadpoles, freshwater snails, fish, frogs |
Worldwide, but human cases seen most frequently in Southeast Asia and in areas where undercooked or raw freshwater snails, clams, and fish are eaten (as of December 27, 2017) |
Heterophyiasis |
Fish |
Egypt, Middle East, Far East (as of January 3, 2018) |
Metagonimiasis |
Fish (cyprinid) |
Far East, Siberia, Manchuria, the Balkan states, Israel, Spain (as of December 29, 2017) |
* Adapted with permission from Tribble D, Wagner KF. Trematode infections. Infectious Disease Practice. 1996; 20:69-73. ** Updated data from the Centers for Disease Control and Prevention (DPDx Laboratory Identification of Parasites of Public Health Concern. Available: http://www.cdc.gov/dpdx/) |
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Infection |
Source |
Clinical Features |
Alaria americana |
Undercooked frog legs |
Disseminated fatal thoracic, gastrointestinal, retroperitoneal, and CNS manifestations; intraocular infections |
Echinostomiasis (16 species) |
Freshwater fish, aquatic plants, clams, snails, mollusks, contact with aquatic birds |
May be asymptomatic; mild abdominal pain, bloating, dyspepsia, diarrhea, eosinophilia |
Fibricola species |
Tadpoles |
Abdominal pain, diarrhea, fever, eosinophilia |
Fasciolopsis species |
Water chestnut, water calthrop, water bamboo, water morning glory lotus and water hyacinth |
May be symptomatic; may be subclinical; gastritis, nausea, diarrhea, eosinophilia; generalized edema in persons with heavy infection burden |
Gastrodiscoides species |
Vegetables, aquatic plants |
Often asymptomatic; may manifest as abdominal pain and diarrhea in severe cases |
Watsonius watsoni |
Water bamboo |
Severe diarrhea |
Fischoederius elongates |
Aquatic plants |
Epigastric pain and vomiting |
Heterophyes species |
Mullets, fish; brackish water |
May be asymptomatic; intestinal mucosal disease, ulcer-related abdominal pain, dyspepsia, nausea, vomiting, diarrhea, weight loss |
Gymnophalloides seoi |
Oysters |
Fever, abdominal pain, anorexia, weight loss, diarrhea, pancreatitis |
Carneophallus brevicaeca |
Shrimp |
Fatal when infection involves CNS and heart |
Brachylaima ruminae |
Poultry, rats |
Abdominal pain, diarrhea |
Metagonimiasis species |
Fish (ayu, golden carp) |
May be asymptomatic; intestinal mucosal disease, ulcer-related abdominal pain, dyspepsia, nausea, vomiting, diarrhea, weight loss |
Nanophyetus salmincola |
Undercooked fish (eg, salmon, trout, steelhead) |
May be symptomatic; mild diarrhea, abdominal pain |
*Adapted from Berger SA, Marr JS. Human Parasitic Diseases Sourcebook. 1st ed. Sudbury, MA: Jones and Bartlett; 2006. |
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